Incorporation of [³⁵S] sulfate into chondroitin sulfates by organized cultures of murine peripheral, sympathetic and central nervous tissues

Abstract

SUMMARYOrganized cultures of mouse dorsal root ganglia (PNS), cerebellum (CNS), and sympathetic chain ganglia (ANS) were exposed to feeding media containing radioactive Na235S04 for 10 day periods beginning either at the onset of myelination or during myelin maintenance. During this period, the used medium was collected at each of three feedings and frozen. Some cultures were frozen and together with the collected medium were analyzed for mucopolysaccharides (MPS) . Sister cultures were fixed in ruthenium red-glutaraldehyde and processed for [ %] radioautography by light microscopy, and cellular localization of MPS by electron microscopy.[W] MPS were isolated from both cultures and medium (by alkali treatment, proteolytic digestion, TCA treatment, and dialysis, followed by precipitation with cetylpyridinium chloride and ethanol). Isolated MPS were analysed by paper chromatography after digestion with chondroitinase-ABC and testicular hyarulonidase.Fifty-five to seventy-five percent of the total sulfated MPS formed in all types of cultures were chondroitin sulfates (Ch-S) A, B, and C,$ chondroitin sulfate A accounting for 50 t o 60% of the total MPS. PNS and ANS exceeded CNS cultures in total sulfated MPS formed by 10 : 1.Qualitatively, CNS cultures produced a higher proportion of ChS-C and a lower proportion of ChS-B compared to PNS and ANS. Rutheniumred positive material covered all types of cell surfaces, collagen fibers, and the surfaces of enveloped axons; the layers of compact myelin and its underlying axon-Schwann cell interface showed no such staining, though it appeared regularly in the external mesaxon.

Myelinogenesis in optic nerve. A morphological, autoradiographic, and biochemical analysis.

“…In addition, there was a linear correlation between diameters of optic nerve axons and the number of lamellae of myelin, with the largest fibers having the greatest numbers of lamellae. Similar data have been obtained on myelin/axon ratios in both the peripheral nervous systems and other parts of the CNS (16)(17)(18) Biochemical Components of Optic Nerve SULFATIDES AND SULFATED GLYCOPRO-TEINS: In the CNS, the important sulfated compounds include sulfatide which is enriched in myelin (39,40), sulfated glycoproteins which are present in small quantities in myelin (35,43), and sulfated mucopolysaccharides which appear to be associated with neurons (31) rather than myelin (52). Sulfate is donated to these compounds by PAPS (6,45).…”

“…AUTORADIOGRAPHIC LOCALIZATION OF LABEL: At 2 days postnatal, the basal lamina of the glia limitans showed a high grain density. This structure, a surface product of astroglia, was thought to be composed in part of sulfated mucopolysaccharides (52). Since astroglia are the major cell type in the first few postnatal days (50,51), it is probable that these cells were active in the sulfation of mucopolysaccharides.…”